Brucella spp. is bacteria that naturally infect a variety of domesticated and wild animals leading to abortions and sterility. These bacteria are also capable of causing debilitating human infections, which often result from human exposure to infected animals and animal products. Additionally, Brucella spp. are considered threats as potential biological weapons. Currently, there is no safe and effective vaccine to protect humans against infection with Brucella, and antibiotic treatment against brucellosis is prone to disease relapse. During the course of an infection, the Brucella reside within immune cells called macrophages where they replicate in a specialized compartment, and the ability of Brucella to survive and replicate within macrophages is essential to their ability to cause disease. We have recently discovered that a transcriptional regulator, which we have named VtlR, is required for the ability of the bacteria to survive and replicate within host cells, and moreover, this regulato is essential for chronic Brucella colonization in an animal model of infection. Preliminary data has led to the hypothesis that VtlR controls the expression of essential genes for Brucella pathogenesis. We have determined that VtlR regulates the expression of a limited subset of three genes putatively encoding small hypothetical proteins. Strikingly, these hypothetical proteins are highly conserved among numerous bacterial species, both pathogenic and beneficial, but there is currently no information regarding the molecular function(s) of these proteins in prokaryotic biology. The objective of this proposal is to define these novel virulence-associated elements in Brucella spp. In turn, these proteins may be targeted in new therapeutic strategies to combat Brucella infections, or to develop a human vaccine against brucellosis. Moreover, these studies will shed light on the role of these novel protein elements in a wide variety of bacteria, and it is possible that these proteins may be exploited as a means of alleviating detrimental infections, as well as enhancing beneficial bacteria-host interactions.